Substantial evidence exists that physiologic fluid shear stress exerts atheroprotective effects in vivo, because atherosclerosis preferentially occurs in areas of disturbed flow or low shear stress, whereas regions with steady laminar flow and physiological shear stress are protected (Gimbrone et al., “Endothelial Dysfunction, Hemodynamic Forces, and Atherogenesis,” Ann NY Acad Sci 902:230-239, Discussion 239-240 (2000); Traub et al., “Laminar Shear Stress: Mechanisms by Which Endothelial Cells Transduce an Atheroprotective Force,” Arterioscler Thromb Vasc Biol 18:677-685 (1998)). Pathogenic features of atherosclerosis are oxidative stress and inflammation characterized by endothelial expression of vascular cell adhesion molecule-1 (“VCAM1”) (Ross, R., “Atherosclerosis—An Inflammatory Disease,” N Eng J Med 340:115-126 (1999)).
Apoptosis signal-regulating kinase 1 (“ASK1”), a mitogen-activated protein kinase kinase kinase (MAPKKK), plays essential roles in cytokine-related signaling and stress-induced apoptosis (Ichijo et al., “Induction of Apoptosis by ASK1, a Mammalian MAPKKK That Activates SAPK/JNK and p38 Signaling Pathways,” Science 275:90-94 (1997)). Through genetic screening for ASK1-binding proteins, Saitoh et al. (“Mammalian Thioredoxin is a Direct Inhibitor of Apoptosis Signal-Regulating Kinase (ASK) 1,” EMBO J 17:2596-2606 (1998)) found that thioredoxin (“TRX”) bound directly to the N-terminus of ASK1 and inhibited ASK1 kinase activity as well as ASK1-dependent apoptosis. TRX is a ubiquitous thiol oxidoreductase that regulates cellular redox status. TRX can protect against oxidative stress-induced cell injury or inflammation directly via antioxidant effects and indirectly by protein-protein interaction with signaling molecules such as ASK1 (Yamawaki et al., “Thioredoxin: A Key Regulator of Cardiovascular Homeostasis,” Circ Res 93:1029-1033 (2003)). TRX also exhibits growth-promoting effects presumably via an increased supply of reducing equivalents for DNA synthesis and activation of transcription factors that regulate cell growth. Thioredoxin interacting protein (“TXNIP,” also termed VDUP1 for vitamin D3-upregulated protein) was originally identified in HL-60 leukemia cells treated with 1,25-dihydroxyvitamin D3 (Chen et al., “Isolation and Characterization of a Novel cDNA from HL-60 Cells Treated with 1,25-Dihydroxyvitamin D-3,” Biochim Biophys Acta 1219:26-32 (1994)). Thereafter, Nishiyama et al. isolated TXNIP as a TRX-binding protein using a yeast two-hybrid system (Nishiyama et al., “Identification of Thioredoxin-Binding Protein-2/Vitamin D(3) Up-Regulated Protein 1 as a Negative Regulator of Thioredoxin Function and Expression,” J Biol Chem 274:21645-21650 (1999)). Biochemical analysis showed that TXNIP inhibited TRX activity by interacting with the catalytic center of TRX (cysteines 32 and 35), suggesting that TXNIP is an endogenous inhibitor of TRX (Nishiyama et al., “Identification of Thioredoxin-Binding Protein-2/Vitamin D(3) Up-Regulated Protein 1 as a Negative Regulator of Thioredoxin Function and Expression,” J Biol Chem 274:21645-21650 (1999); Junn et al., “Vitamin D3 Up-Regulated Protein 1 Mediates Oxidative Stress via Suppressing the Thioredoxin Function,” J Immunol 164:6287-6295 (2000)).
There is accumulating evidence that TXNIP plays a pivotal role in cardiovascular disorders, functioning as a sensor for biomechanical and oxidative stress. Schulze et al. recently reported that hyperglycemia in vascular smooth muscle increased oxidative stress by inducing TXNIP and inhibiting the anti-oxidant function of TRX (Schulze et al., “Hyperglycemia Promotes Oxidative Stress Through Inhibition of Thioredoxin Function by Thioredoxin-Interacting Protein,” J Biol Chem 279:30369-30374 (2004)). They also showed that diabetic animals exhibited increased vascular expression of TXNIP. Wang et al. recently demonstrated in cardiomyocytes that mechanical strain suppressed TXNIP expression followed by increases in TRX activity (Wang et al., “Vitamin D(3)-Up-Regulated Protein-1 is a Stress-Responsive Gene That Regulates Cardiomyocyte Viability Through Interaction with Thioredoxin,” J Biol Chem 277:26496-26500 (2002). It has also been reported by Yoshioka et al. that TXNIP expression is decreased in pressure-overload cardiac hypertrophy followed by TRX-induced stimulation of cardiac cell growth (Yoshioka et al., “Thioredoxin-Interacting Protein Controls Cardiac Hypertrophy Through Regulation of Thioredoxin Activity,” Circulation 109:2581-2586 (2004)).
It would be desirable, therefore, to identify whether inhibition of TXNIP in endothelial cells can have an atheroprotective effect via increased TRX activity, and decreased activity of JNK, p38, and VCAM1 expression. The present invention is directed to overcoming the above-identified deficiencies in the art.